X-ray absorption spectroscopy at the molybdenum and sulfur K-edges has been used to probe the active site of wild-type and cysteine 207 . serine mutant human sulfite oxidases. The active site structures of the oxidized (resting) Mo(VI) enzymes have been compared. The wild-type enzyme has been shown to possess two Mo=O ligands at 1.71 E and three Mo-S ligands at 2.41 E. The mutant enzyme possesses a novel tri-oxo molybdenum site with three Mo=O ligands at 1.74 E and two Mo-S ligands at 2.47 E. This work shows that cysteine 207 is a ligand of molybdenum in wild-type human sulfite oxidase, and that in the mutant the Mo is ligated to an extra oxo group rather than to the hydroxyl of the substituant serine 207. This work has been published: J. Am. Chem. Soc. 1996, 118, 8588-8592